The main objective of the NAMDC is the precise characterization of mitochondrial patients as a foundation for clinical investigation and treatment trials. Capitalizing upon the patient cohort and the in vivo imaging resources provided by NAMDC and UT Southwestern, our overarching goal is the quantitative analysis of human muscle and brain metabolism in a non-invasive, focused manner using ultra high-field (7T) nuclear magnetic resonance (NMR) spectroscopy (MRS). This pilot project represents the translation, for the first time, of extensive work we have conducted in animals and man to the evaluation of mitochondrial dysfunction in vivo. This Pilot project's goal is to utilize information-rich MRS spectra arising from metabolites whose turnover depends upon mitochondrial function to report dysfunction, Among all MRS-observable metabolites, physiological considerations guide the selection of MRS target analytes: For muscle, we will determine intracellular lipid abundance because it represents, for the purposes of MRS, a stable metabolic pool relative to other biochemical pools subject to rapid physiological blood-muscle abundance fluctuations. For the brain, we will take advantage of the robust production of glutamate by neuronal mitochondria relative to glia (which generate predominantly glutamine) and of the differential impact of mitochondrial diseases on neurons relative to glia, to evaluate the ratio between these two substrates as an indicator of mitochondrial dysfunction severity. The project will include a pilot group of patients afflicted by the mitochondrial disease MELAS (mitochondrial encephalomyopathy, lactic acidosis and stroke-tike episodes), together with proper control subjects, characterized in sufficient detail to conduct hypothesis testing. Patients will be assessed by scored clinical criteria, clinical blood metabolic indicators and genotyping and by MRS to investigate two specific aims relevant to mitochondrial function in muscle and in brain. Aim 1: To test whether intramyocellular lipid (IMCL) accumulation, the result of impaired fat oxidation in muscle mitochondria, correlates with clinical disease severity and insulin resistance; and Aim 2: To test the hypothesis that reduced brain glutamate turnover relative to glutamine, which is dependent upon neuronal relative to glial mitochondrial tricarboxylic acid (TCA) cycle activity, is a characteristic of the disease state. Our preliminary results illustrate that the significance of the 7T MRS approach transcends its innovation: (1) IMCL abundance is increased in MELAS, including patients additionally manifesting diabetes; (2) brain glutamate/ glutamine turnover determined by ex vivo NMR of brain samples is decreased in biological settings where neuronal mass is decreased relative to glial mass such as normal pig white matter or human glioblastoma; and (3) precise in vivo MRS determination of human brain glutamate/ glutamine ratio can be achieved by multiplet MRS. It is anticipated that the approach will also be widely applicable to other metabolic diseases and states associated with mitochondrial dysfunction.